Molecular Mechanisms of Myofibril Assembly in Heart

Cell Structure and Function

Elisabeth Ehler¹  Jean-Claude Perriard¹  David Helfman¹  Daniel Auerbach¹  Stefan Bantle¹  Barbara Rothen-Ruthishauser¹  Martin Leu¹ 

[1] Institute for Cell Biology, Swiss Federal Institute of Technology

关键词: Thick filament assembly;    M-band;    myomesin;    epitope tagging;    green fluorescent protein;    confocal microscopy;   

DOI  :  10.1247/csf.22.139

学科分类：分子生物学，细胞生物学和基因

来源: Japan Society for Cell Biology

【 摘 要 】

References(27)Cited-By(14)We investigated the assembly of the first sarcomeres in chicken embryos by conf ocal microscopy of immunofluorescently stained whole mount rudiments of early chicken hearts isolated around the onset of beating. In embryos with merely 9 somites, myomesin was found to be present in a cross striated pattern, indicating that myomesin is expressed rather early during development. RNA studies confirmed these findings and RT-PCR revealed the presence of myomesin mRNA already in the 7 somite embryo. The expression of myomesin mRNA coding for the skeletal isoform preceded the heart specific transcript. In the adult heart, however, only the heart isoform was detectable. The interaction of myomesin domains with the sarcomere was investigated by transf ection of epitope tagged constructs into cultured cardiomyocytes. The second domain of myomesin, an immunoglobulin-like domain, was found to specifically bind to the M-band region of chicken cardiomyocytes. All constructs containing this domain also showed M-band localization. Additionally, constructs consisting of either the second domain of myomesin or the myosin light chain isoform MLC 3f fused to the green fluorescent protein (eGFP) were expressed in rat cardiomyocytes and were found to be distributed in the same manner as the expression constructs tagged only with the much shorter VSV epitope. Transfected cells did not show any alteration in beating activity and no alteration of myofibrillar structure, as judged by simultaneous staining for the Z-disc protein α-actinin and other sarcomeric markers. Apparently, the addition of eGFP did not disturb the assembly properties or the function of the two proteins and therefore allowed the easy visualization of assembly and contraction processes directly in the living cardiomyocyte.

【 授权许可】

Unknown